PROJECT TITLE: The Role of Glucocerebrosidase and Chaperone-mediated Autophagy in Parkinson’s Disease

Investigators/Authors: Sheng-Han Kuo, M.D., Columbia University

Objective: In this proposal, we will investigate whether the disease-causing glucocerebrosidase (GBA) mutations can perturb a major pathway for a-synuclein degradation, chaperone-mediated autophagy (CMA) and subsequently lead to a-synuclein accumulations and neurodegeneration, two hallmarks of Parkinson’s disease (PD) pathology.

Background: GBA mutations are the most common genetic risk factors for PD and the majority of PD patient brains have a-synuclein aggregation, called Lewy bodies. How GBA mutation cause a-synuclein accumulation remains uncertain. We specifically investigate the effect of mutant GBAs on the a-synuclein degradation pathway, CMA.

Methods/Design: We will introduce either wild type GBA or two disease-related mutant GBAs by lentivirus to the primary mouse dopaminergic neuronal cultures. We will assess whether expressing the exogenous mutant GBAs can interfere with 1) CMA activity assessed by a novel CMA reporter, 2) a-synuclein protein levels, and 3) neuronal death. We will examine whether toxic effects of GBA mutations act through a-synuclein by introducing mutant GBAs into primary dopaminergic neuronal cultures that do not have a-synuclein. If toxic effects of GBA mutations are mediated via a-synuclein, we can expect that genetic elimination of a-synuclein can rescue neuronal toxicity of mutant GBA. Finally, we will investigate the role of CMA in this pathway. We will delete the essential part of mutant GBAs that is required to interfere with CMA. We expect that this manipulation can eliminate the toxic effects of mutant GBAs on a-synuclein accumulation and neuronal death, highlighting the importance of CMA. This study will comprehensively examine the relationship between GBA, CMA, and a-synuclein.

Relevance to Parkinson’s disease: We will investigate the mechanism behind the most common form of genetic PD with the GBA mutations and how these mutations can lead to the PD pathology, specifically focusing on CMA. The current research proposal will help us to have a better understanding of the PD disease mechanism and the intervention on CMA might be a promising strategy to treat PD.

Objectives: 1) To determine the efficacy of a MHT-mediated exercise program to improve physical activity six months after randomization in persons with PD compared to a controlled condition without MHT; 2) compare the percentage of days within that period that subjects and controls exceed the minimum threshold index indicating a sedentary lifestyle; 3) determine whether a MHT-mediate exercise program improves motor function, non-motor function, quality of life, exercise self-efficacy and outcome expectations in persons with PD compared to controls.

Background: PD causes significant function decline resulting in greater disability, but can be improved through exercise. People with PD have long lifespans so exercise is essential in improving their function, and because indefinite PT is not realistic, interventions using MHT allow therapists to stay connected over time to meet patients’ changing needs. Innovative approaches using MHT may improve outcomes, however, the effectiveness of different approaches to improve function to reduce disability is unknown. The purpose of this study is to compare the effectiveness of two interventions to improve function and health-related quality of life in people with PD.

Methods/Design: Fifty persons after a baseline assessment are randomly allocated into one of two home exercise conditions for six months with one in-person instructional session and a final assessment. One group receives a home exercise program in written format to continue independently. The other group has the exercise program delivered via videos on a computer table that allow the PT to remotely monitor progress and modify the program to meet changing needs. The long-term objective is to determine the most efficient and effective way to improve function. The primary outcome for the study is the change in physical activity level from the baseline to the six-month assessment measured by recording step counts across seven days before and seven days after the six-month intervention. Other outcomes include disease severity, quality of life, walking ability, balance, function related to non-motor symptoms and confidence in ability to exercise.

Relevance to Parkinson’s disease: The importance of physical activity for persons with Parkinson’s disease has been drastically undervalued. A sedentary lifestyle hastens disability. Exercise prescription is not part of the standard clinical management of persons with the disease. Effective strategies to increase physical activity are essential in reducing disability, improving physical function and enhancing quality of life. If this approach is successful, the standard clinical management of persons with PD could be impacted. This approach is easily accessible to most, generalizable to many, and conducive to widespread dissemination.

Objective: Lewy Bodies (LBs) and Lewy Neurites (LNs) are among the primary hallmarks of Parkinson’s disease. These are protein aggregates found in the cell bodies and axons of neurons which are mostly composed of the protein, a-synuclein (a-syn). At the University of Pennsylvania, a novel model was developed in which, for the first time, we can induce formation of LB and LN-like aggregates in neurons in culture. In this model, pathologic fibrils of a-syn are formed in a test tube and added directly to neurons in a culture dish. These “pre-formed” fibrils, lead to the recruitment of normal a-syn expressed in the neurons into abnormal aggregates. Over time, these aggregates lead to neuron death. This model allows us to understand how these abnormal aggregates form, their impact on neuron function and, ultimately, identify therapeutic targets to prevent their formation.

Background: a-Syn and LRRK2 are 2 autosomal dominant causes of Parkinson’s disease (PD) and among the top genetic susceptibility loci identified in genome wide association studies. In addition, LRRK2 has recently emerged as a therapeutic target for PD and potent and specific inhibitors of LRRK2 have already been developed. The familial linked LRRK2-G2019S mutation increases the activity of the protein which has been associated with neuron death. We will determine if the LRRK2-G2019S mutant accelerates a-syn pathology and conversely, if LRRK2 inhibitors prevent the formation of abnormal LBs and LNs.

Methods/Design: We will use antibodies that recognize the abnormal form of a-syn and imaging with a confocal microscope (a microscope that is able to filter out the out-of-focus light from above and below the point of focus in the object) to visualize its formation over time. Biochemical methods will be used to quantify conversion of a-syn from its normal, soluble form to an abnormal form that becomes insoluble in detergent as it is converted into LB- and LN- like aggregates. We will determine if specific inhibitors of LRRK2 activity will prevent this conversion of a-syn into its abnormal form. Neuron cultures from the transgenic mice expressing LRRK2-G2019S will be used to determine if this familial mutant causes a-syn aggregates to form faster, consequently accelerating cell death.

Relevance to Parkinson’s disease: These studies will begin to determine if LRRK2 inhibitors can prevent the formation of neuronal pathologic aggregates and enhance neuron survival reducing or eliminating the threat of cell death.